Tetrahymena is a ciliated eukaryotic unicellular organism belonging to the regnum of Protozoa and bearing two nuclei, a transcriptionally silent, diploid germline micronucleus (MIC) and a transcriptionally active, polyploid somatic macronucleus (MAC). In 1923, when Nobel Laureate Andre Lwoff succeeded in growing Tetrahymena in pure culture, the basis for exploiting this alveolate as a model organism was laid. Milestone discoveries made in Tetrahymena are the discovery of dynein motors, telomeres, RNA-mediated catalysis, telomerase and the function of histone acetyltransferases in transcription regulation. Within the last decades molecular biological techniques have been developed to alter Tetrahymena's genome and proteome: DNA transfection methods comprise inter alia microinjection into the MAC by electroporation and biolistic bombardment of MIC and MAC. Episomal plasmids based on an rDNA-replicon are available, as well as knock-out/-in techniques based on homologous recombination. On protein level, heterologous expression of related species has been performed and also endogenous proteins were silenced by a novel antisense-ribosome-technique. The advantages of using Tetrahymena in biotechnological applications include fast growth, high biomass, fermentation in ordinary bacterial/yeast equipment, up-scalability as well as existence of cheap and chemically defined media.
So far, only a few markers that can be used in Tetrahymena have been described: ribosomal point mutation mediated resistances, a plasmid based neomycin resistance and a complicated beta-tubulin selection marker making use of an inducible promotor in combination with mutated tubulins being resistant/sensitive to the mitotic drug taxol1. Yet no true auxotrophic marker is available that permits selection without the use of antibiotics or drugs. This is where the present invention applies.